Reference to any prior art in the specification is not and should not be taken as an acknowledgement or any form of suggestion that this prior art forms part of the common general knowledge in Australia or any other jurisdiction or that this prior art could reasonably be expected to be ascertained, understood and regarded as relevant by a person skilled in the art.
Purinergic (P2X) receptors are ATP-gated cation-selective channels. Each receptor is made up of three protein subunits or monomers. To date seven separate genes encoding P2X monomers have been identified: P2X1, P2X2, P2X3, P2X4, P2X5, P2X6, P2X7.
P2X7 receptors are of particular interest as the expression of these receptors is understood to be limited to cells having potential to undergo programmed cell death, such as thymocytes, dendritic cells, lymphocytes, macrophages and monocytes. There is some expression of P2X7 receptors in normal homeostasis, such as on erythrocytes.
Interestingly, a P2X7 receptor containing one or more monomers having a cis isomerisation at Pro210 (according to SEQ ID No: 1 in FIG. 1) and which is devoid of ATP binding function has been found on cells that are understood to be unable to undergo programmed cell death, such as preneoplastic cells and neoplastic cells. This isoform of the receptor has been referred to as a “non functional” receptor.
Antibodies generated from immunisation with a peptide including Pro210 in cis bind to non functional P2X7 receptors. However, they do not bind to P2X7 receptors capable of binding ATP. Accordingly, these antibodies are useful for selectively detecting many forms of carcinoma and haemopoietic cancers and to treatment of some of these conditions.
WO02/057306A1 and WO03/020762A1 both discuss a probe for distinguishing between functional P2X7 receptors and non functional P2X7 receptors in the form of a monoclonal antibody.
Monoclonal antisera have certain serological characteristics not found in polysera that make monoclonal antisera particularly valuable reagents for use in research, diagnosis and therapy. Key amongst these is that monoclonal antibodies generally have an affinity for antigen that is higher than the affinity of most of specificities found in a polysera.
To date it has been very difficult to obtain a hybridoma that generates useful amounts of monoclonal antibody against non functional P2X7 receptors as expressed on live cells, and in particular, monoclonal antibodies that can be used in a range of diagnostic and therapeutic applications. Furthermore, the inventors are not aware of any monoclonal antibodies that have been generated against functional P2X7 receptors on live cells. There is a need for such reagents, particularly for new antibodies capable of discriminating between ATP and non-ATP binding P2X7 receptors on live cells.
Further, as far as the inventors are aware, anti-P2X7 antibodies generally do not discriminate between P2X7 monomers and the trimeric P2X7 receptor formed from these monomers. Antibodies that bind to the trimeric receptor but not to P2X7 monomers would be advantageous for staging a cancer, given that the trimeric receptor is particularly found on advanced neoplastic tissue.